A functional and gene regulatory analysis of the effects of interferon gamma treatment on the dendritic cell response to Aspergillus fumigatus infection in the setting of calcineurin inhibition

Abstract

Invasive pulmonary aspergillosis is a lethal opportunistic fungal infection with high mortality in lung transplant recipients on calcineurin inhibitors. An understanding of the host immune response is critical to improving clinical outcomes. In this thesis I characterize the function of dendritic cells, which bridge innate and adaptive immunity, in response to Aspergillus fumigatus during calcineurin inhibition, and ascertain a potential role of interferon gamma in augmenting the host response. I examined the effects of calcineurin inhibitors on the key dendritic cells pathogen responses of phagocytosis, maturation and cytokine secretion, using a monocyte derived dendritic cell model. I used a mixed leucocyte reaction to analyze T cell stimulatory capacity of dendritic cells, and then introduced macrophages to assess the effect of calcineurin inhibitors and interferon gamma on macrophage fungal killing by dendritic cells indirectly via T cells. Calcineurin inhibitors impaired dendritic cell maturation in response to A. fumigatus, impaired CD4 Th1 polarization and macrophage fungal killing. Interferon gamma restored maturation, promoted the Th1 phenotype and improved macrophage phagocytosis via a T cell dependent effect on dendritic cells. I evaluated regulatory transcriptomic pathways in dendritic cells and macrophages by RNA-Seq, finding a divergence of transcriptional pathways of myeloid cells commensurate with their innate functions. Interferon gamma elicited gene expression profiles in dendritic cells suggestive of activation of non calcineurin-NFAT immune pathways. I investigated regulation of gene expression in dendritic cells by identifying active chromatin elements by histone modification ChIP-Seq. This revealed dominant control by an interferon gamma-induced regulatory switch from STAT3 to STAT1 transcription factor binding in the control of A. fumigatus infection during calcineurin inhibitor treatment. These findings reveal mechanisms by which interferon gamma treatment augments the dendritic cell mediated immune response to A. fumigatus infection during calcineurin inhibitor treatment, and provides support for its clinical use as immunoadjuvant therapy in this patient population.